The neural crest is a transient population of cells, named because it arises on the "crest" of the closing neural tube. Neural crest cells emigrate from the neural tube, migrate along precise pathways and finally localize in characteristic sites in the embryo to form the ganglia of the peripheral nervous system, as well as melanocytes and the craniofacial skeleton. They form the autonomic nervous system that innervates numerous organs such as the gut, kidneys and pancreas. Correct migration to and innervation of these targets is essential for proper body function and homeostasis. During the previous grant period, we showed that the dominant guidance cues for neural crest migration may be inhibitory: interactions between Eph receptors on neural crest cells and ephrins in the caudal somite results in a segmentally restricted migratory pattern in the trunk. In addition, Slit chemorepellants expressed prevent trunk neural crest cells from invading the gut (deBellard et al., 2003). Using a genomics approach, we have analyzed the repertoire of genes expressed by premigratory neural crest cells. This has provided us with numerous new candidate genes that may function in the migratory process. Genes identified in neural folds include Slit, laminin a5, as well as a novel chemokine. The proposed experiments will characterize the function of these candidate genes in regulating important events in neural crest migration and other developmental processes. We will use both in vivo and in vitro experiments combining gain-of-function and loss-of-function approaches to examine: 1. The role of the dual inhibitory and migration-stimulating activity of Slit in the migration of trunk and cranial neural crest cells. 2. The function of the a5 subunit of laminin in early neural crest development. 3. The function of a novel chemokine in neural crest migration. 4. The function of this novel chemokine in limb development.